This application proposes detailed analysis of the genetics and physiology of the IgA proteases of Neisseria gonorrhoeae. These enzymes, which specifically proteolyze human IgAl, have been linked to virulence in this and other bacterial species, although their exact role in pathogenesis has not been defined. Unlike many virulence factors such as capsules or pili, these enzymes, which are secreted to the extracellular medium by these gram-negative bacteria, are maintained in Neisseria in the absence of selective pressure and their role in the physiology of the cell remains unclear. IgA proteases of two distinct specificities are produced by N. gonorrhoeae, and enzyme type is apparently linked with nutritional auxotype and/or protein 1 serogroup. Genes specifying types 1 and 2 gonococcal IgA proteases will be cloned into E. coli K-12, and the structures of the cloned genes characterized and compared using restriction enzymes, specific mutations, and DNA sequence analysis of the regulatory regions and the sequences encoding the amino-terminal section of the polypeptide. Mutant genes, especially those encoding truncated proteins, and their products will be analyzed to localize regions of the protein involved in active, binding, and antigenic sites, and in protein secretion. Selected mutations will be transformed into N. gonorrhoeae cells to construct defined IgA protease negative strains. Isogenic protease positive and negative strains will be used to examine the role of these enzymes in gonococcal physiology, and in model systems of gonococcal infection. Linkage of IgA protease genes to genes responsible for nutritional requirements, outer membrane proteins, serum resistance, and antibiotic sensitivity will be assessed by cotransformation and by chromosome walk techniques. The objectives of this research are to clarify the specific role of IgA proteases in gonococcal physiology and in the infective process and to extend our knowledge of the pathogenesis of gonococcal infection and the genetic control of virulence in N. gonorrhoeae.